摘要
本研究以山羊睾丸组织为材料,用RT-PCR法成功克隆了山羊DAZL基因的cDNA序列950 bp,测序和生物信息学分析表明,其含有885 bp的完整开放阅读框(ORF),编码295个氨基酸,与牛的氨基酸序列同源性为97.97%,编码产物含有典型的RNA识别基序RRM和DAZ重复基序;将山羊DAZL基因亚克隆至表达载体pEGFP-C1上,构建了山羊DAZL基因真核表达载体pEGFP-DAZL,采用脂质体法将其瞬时转染至山羊骨髓间充质干细胞(BMSC)中,通过荧光显微镜观察确定重组质粒在山羊BMSC内的表达和定位。
DAZL（Deleted in Azoospermia-Like）,a germ cell-specific gene,has shown to regulate meiosis during spermatogenesis and play important roles in germ cell development and differentiation.In this study,goat DAZL gene coding region 950 bp was successfully amplified by RT-PCR from goat testis,and the sequence analysis showed that the CDS region included 885 nucleotides（ORF）,encoding for 295 amino acids,and containing typical RRM domain and DAZ repeat motif.The amino acid similarity of DAZL gene between goat and bovine was 97.97%.The DAZL gene was then subcloned into eukaryotic express vector pEGFP-C1 to generate recombinant expression vector pEGFP-DAZL.Furthermore,we transfected pEGFP-DAZL plasmid into goat bone marrow mesenchymal stem cells（BMSC）by liposome,and its expression was identified by observation of fluorescence microscopy and RT-PCR.
出处
《南京农业大学学报》
CAS
CSCD
北大核心
2012年第6期104-110,共7页
Journal of Nanjing Agricultural University
基金
国家自然科学基金青年基金项目(31201802)
转基因生物新品种培育重大专项(2011ZX08008-003)
南京农业大学青年科技创新基金项目(KJ2010012)
关键词
山羊
DAZL基因
序列分析
真核表达
骨髓间充质干细胞
goat
DAZL gene
sequence analysis
eukaryotic expression
bone marrow mesenchymal stem cells（BMSC）
